Sepsis is a major problem in medicine, and jeopardizes the health of nearly 800,000 persons in the United States annually, killing >200,000. Recent knowledge from work supported during the prior years of this grant reveals that neural circuits regulate cytokine production to prevent potentially damaging inflammation. This has enabled development of experimental drugs to modify neural-immune crosstalk, and prevent lethal inflammation in sepsis. A prototypical vagus nerve circuit, the inflammatory reflex, inhibit cytokine production in spleen through a mechanism requiring acetylcholine signaling through 7 nAChR expressed on cytokine- producing macrophages. Nerve fibers in spleen lack the enzymatic machinery necessary for acetylcholine production, but we have recently identified an acetylcholine-producing T cell population that is required for the cytokine inhibiting mechanism of the inflammatory reflex. At this writing, these data are in press in Science, giving evidence of the innovation and significance of the findings. We are requesting funding to focus on the next unanswered questions. Here, we hypothesize that acetylcholine-producing T cells play a pivotal role in the regulation of cytokine production during severe sepsis, and that targeting these cells in this neural to immune pathway will be useful in the development of experimental therapeutics for severe sepsis. This hypothesis will be addressed in the following three Specific Aims: Specific Aim 1. To study the role of acetylcholine producing T cells in sepsis survival. Specific Aim 2. To study the molecular mechanisms underlying T cell acetylcholine production. Specific Aim 3. To develop experimental therapeutics that target T cell acetylcholine release. We propose to utilize an innovative strategy to selectively isolate acetylcholine synthesizing T cells and then knock out ChAT expression to study the specific requirement for ChAT expression to confer protection. A successful completion of this research will provide significant data that can be used to modulate this neural circuitry to develop therapeutic modalities for the prevention and treatment of sepsis/septic shock. PUBLIC HEALTH RELEVANCE: Severe sepsis is a major public health problem, because it is the leading cause of death in hospitalized patients in the United States, and one of the ten leading causes of death in the developed world. Recent evidence indicates that neural circuits regulate cytokine production to prevent potentially damaging inflammation. The studies proposed here will provide significant data that can be used to modulate this neural circuitry to develop therapeutic modalities for the prevention and treatment of sepsis/septic shock.